Stress-induced c-Fos Expression Research Articles

Dominance status modulates activity in medial amygdala cells with projections to the bed nucleus of the stria terminalis

The medial amygdala (MeA) controls several types of social behavior via its projections to other limbic regions. Cells in the posterior dorsal and posterior ventral medial amygdala (MePD and MePV, respectively) project to the bed nucleus of the stria terminalis (BNST) and these pathways respond to chemosensory cues and regulate aggressive and defensive behavior. Because the BNST is also essential for the display of stress-induced anxiety, a MePD/MePV-BNST pathway may modulate both aggression and responses to stress. In this study we tested the hypothesis that dominant animals would show greater neural activity than subordinates in BNST-projecting MePD and MePV cells after winning a dominance encounter as well as after losing a social defeat encounter. We created dominance relationships in male and female Syrian hamsters (Mesocricetus auratus), used cholera toxin b (CTB) as a retrograde tracer to label BNST-projecting cells, and collected brains for c-Fos staining in the MePD and MePV. We found that c-Fos immunoreactivity in the MePD and MePV was positively associated with aggression in males, but not in females. Also, dominant males showed a greater proportion of c-Fos+ /CTB+ double-labeled cells compared to their same-sex subordinate counterparts. Another set of animals received social defeat stress after acquiring a dominant or subordinate social status and we stained for stress-induced c-Fos expression in the MePD and MePV. We found that dominant males showed a greater proportion of c-Fos+ /CTB+ double-labeled cells in the MePD after social defeat stress compared to subordinates. Also, dominants showed a longer latency to submit during social defeat than subordinates. Further, in males, latency to submit was positively associated with the proportion of c-Fos+ /CTB+ double-labeled cells in the MePD and MePV. These findings indicate that social dominance increases neural activity in BNST-projecting MePD and MePV cells and activity in this pathway is also associated with proactive responses during social defeat stress. In sum, activity in a MePD/MePV-BNST pathway contributes to status-dependent differences in stress coping responses and may underlie experience-dependent changes in stress resilience.

Effects of footshock stress on social behavior and neuronal activation in the medial prefrontal cortex and amygdala of male and female mice.

Social behavior is complex and fundamental, and its deficits are common pathological features for several psychiatric disorders including anxiety, depression, and posttraumatic stress disorder. Acute stress may have a negative impact on social behavior, and these effects can vary based on sex. The aim of this study was to explore the effect of acute footshock stress, using analogous parameters to those commonly used in fear conditioning assays, on the sociability of male and female C57BL/6J mice in a standard social approach test. Animals were divided into two main groups of footshock stress (22 male, 24 female) and context exposed control (23 male and 22 female). Each group had mice that were treated intraperitoneally with either the benzodiazepine-alprazolam (control: 10 male, 10 female; stress: 11 male, 11 female), or vehicle (control: 13 male, 12 female; stress: 11 male, 13 female). In all groups, neuronal activation during social approach was assessed using immunohistochemistry against the immediate early gene product cFos. Although footshock stress did not significantly alter sociability or latency to approach a social stimulus, it did increase defensive tail-rattling behavior specifically in males (p = 0.0022). This stress-induced increase in tail-rattling was alleviated by alprazolam (p = 0.03), yet alprazolam had no effect on female tail-rattling behavior in the stress group. Alprazolam lowered cFos expression in the medial prefrontal cortex (p = 0.001 infralimbic area, p = 0.02 prelimbic area), and social approach induced sex-dependent differences in cFos activation in the ventromedial intercalated cell clusters (p = 0.04). Social approach following stress-induced cFos expression was positively correlated with latency to approach and negatively correlated with sociability in the prelimbic area and multiple amygdala subregions (all p < 0.05). Collectively, our results suggest that acute footshock stress induces sex-dependent alterations in defensiveness and differential patterns of cFos activation during social approach.

Early-life Stress Modifies the Reactivity of Neurons in the Ventral Tegmental Area and Lateral Hypothalamus to Acute Stress in Female Rats

Early-life stress (ELS) has long-term consequences, including an increased risk for drug abuse and psychiatric disorders later in life, which is higher in women than in men. The consequences of ELS include heightened sensitivity to stressful events. Here, we hypothesized that ELS changes the stress sensitivity of dopaminergic (DAergic) neurons in the ventral tegmental area (VTA) and orexin (OXA) neurons in the lateral hypothalamus (LH), that are crucial for the control of motivated behaviors. We combined morphological and immunohistochemical approaches to investigate the effect of maternal separation (MS), a rodent model of the ELS, on the expression of c-Fos protein in putative DAergic and non-DAergic VTA and LH OXA neurons, as well as on dendritic spine density and morphology in the VTA of adult female rats. We showed that MS increased basal and acute restraint stress-induced c-Fos expression in putative DAergic neurons, specifically in the dorsomedial VTA, an area implicated in responsiveness to aversive stimuli. Conversely, restraint-induced increase in c-Fos expression in non-DAergic dorsolateral VTA neurons was dampened by MS. Furthermore, an increase in spine head diameter of VTA neurons and a concurrent decrease in dendritic spine density in dorsal VTA were observed. We also showed that MS changed the stress sensitivity of OXA neurons selectively in the dorsomedial hypothalamus (DMH), which is implicated in arousal and the stress response. These findings show the long-lasting consequences of ELS and indicate the selective, regional sensitivity of structures involved in the control of arousal, motivational behaviors and the stress response to ELS.

Social defeat stress-specific increase in c-Fos expression in the extended amygdala in mice: Involvement of dopamine D1 receptor in the medial prefrontal cortex

We recently reported that dopamine D1 receptor in the medial prefrontal cortex (mPFC) is activated by subthreshold social defeat stress and suppresses the induction of depressive-like behavior in mice. However, which mPFC projection(s) mediates this antidepressant-like effect remains poorly understood. Here we show that social defeat stress specifically increased c-Fos expression, a marker for neuronal activity, in distinct brain regions involved in emotional regulation, relative to novelty-induced exploration. Among these brain areas, D1 knockdown in the mPFC decreased social defeat stress-induced c-Fos expression in the interstitial nucleus of the posterior limb of the anterior commissure (IPAC), a subregion of the extended amygdala. Using retrograde adeno-associated virus vectors and transgenic mice expressing Cre recombinase under the D1 promoter, we also found that D1-expressing deep-layer pyramidal neurons in the mPFC send direct projections to the IPAC. These findings indicate that social defeat stress specifically activates neurons in distinct brain areas, among which the IPAC is regulated by dopamine D1 receptor in the mPFC perhaps through direct projections. Thus, this study provides hints toward identifying neural circuits that underlie antidepressant-like effects of stress-induced dopamine D1 receptor signaling in the mPFC.

Chronic stress produces enduring sex- and region-specific alterations in novel stress-induced c-Fos expression

Prolonged or repeated exposure to stress increases risk for a variety of psychological disorders, many of which are marked by dysfunction of corticolimbic brain regions. Notably, women are more likely than men to be diagnosed with these disorders, especially when onset of symptoms follows stressful life events. Using rodent models, investigators have recently begun to elucidate sex-specific changes in the brain and behavior that occur immediately following chronic stress. However, little is known regarding the lasting sequelae of chronic stress, as well as how potential changes may impact responsivity to future stressors. We recently demonstrated that male and female rats show different patterns of dendritic reorganization in medial prefrontal cortex in the days following chronic stress. Here, we examined the immediate and lasting effects of chronic restraint stress (CRS; 3 h/day, 10 days) on neuronal activation, across several corticolimbic brain regions, induced by novel acute stress exposure. Chronically stressed male and female rats were exposed to acute elevated platform stress (EPS) either 1 (CRS-EPS) or 7 (CRS-Rest-EPS) days after CRS. Compared to rats exposed to EPS only, significant reductions in acute stress-induced c-Fos expression were observed in the medial prefrontal cortex, hippocampus, and paraventricular nucleus of the hypothalamus (PVN) in CRS-EPS male rats, some of which persisted to 7 days post-stress. In contrast, we found little modulation of novel stress-induced c-Fos expression in CRS-EPS female rats. However, CRS-Rest-EPS female rats exhibited a significant enhancement of acute stress-induced neuronal activity in the PVN. Together, these data show that prior chronic stress produces sex- and region-specific alterations in novel stress-induced neuronal activation, which are dependent on the presence or absence of a rest period following chronic stress. These findings suggest that the post-stress rest period may give rise to sex-specific neuroadaptations to stress, which may underlie sex differences in stress susceptibility versus resilience.

A Group of Descending Glutamatergic Neurons Activated by Stress in Corticolimbic Regions Project to the Nucleus Accumbens

The nucleus accumbens (NAc) is the major component of the ventral striatum that regulates stress-induced depression. The NAc receives dopaminergic inputs from the ventral tegmental area (VTA), and the role of VTA-NAc neurons in stress response has been recently characterized. The NAc also receives glutamatergic inputs from various forebrain structures including the prelimbic cortex (PL), basolateral amygdala (BLA), and ventral hippocampus (vHIP), whereas the role of those glutamatergic afferents in stress response remains underscored. In the present study, we investigated the extent to which descending glutamatergic neurons activated by stress in the PL, BLA, and vHIP project to the NAc. To specifically label the input neurons into the NAc, fluorescent-tagged cholera toxin subunit B (CTB), which can be used as a retrograde neuronal tracer, was injected into the NAc. After two weeks, the mice were placed under restraint for 1 h. Subsequent histological analyses indicated that CTB-positive cells were detected in 170~680 cells/mm2 in the PL, BLA, and vHIP, and those CTB-positive cells were mostly glutamatergic. In the PL, BLA, and vHIP regions analyzed, stress-induced c-Fos expression was found in 20~100 cells/mm2. Among the CTB-positive cells, 2.6% in the PL, 4.2% in the BLA, and 1.1% in the vHIP were co-labeled by c-Fos, whereas among c-Fos-positive cells, 7.7% in the PL, 19.8% in the BLA, and 8.5% in the vHIP were co-labeled with CTB. These results suggest that the NAc receives a significant but differing proportion of glutamatergic inputs from the PL, BLA, and vHIP in stress response.

Both Basal and Acute Restraint Stress-Induced c-Fos Expression Is Influenced by Age in the Extended Amygdala and Brainstem Stress Centers in Male Rats.

The hypothalamus-pituitary-adrenal axis (HPA) is the main regulator of the stress response. The key of the HPA is the parvocellular paraventricular nucleus of the hypothalamus (pPVN) controlled by higher-order limbic stress centers. The reactivity of the HPA axis is considered to be a function of age, but to date, little is known about the background of this age-dependency. Sporadic literature data suggest that the stress sensitivity as assessed by semi-quantitation of the neuronal activity marker c-Fos may also be influenced by age. Here, we aimed at investigating the HPA activity and c-Fos immunoreactivity 2 h after the beginning of a single 60 min acute restraint stress in eight age groups of male Wistar rats. We hypothesized that the function of the HPA axis (i.e., pPVN c-Fos and blood corticosterone (CORT) level), the neuronal activity of nine stress-related limbic areas (i.e., magnocellular PVN (mPVN), medial (MeA), central (CeA), basolateral nuclei of the amygdala, the oval (ovBNST), dorsolateral (dlBNST), dorsomedial (dmBNST), ventral and fusiform (fuBNST) divisions of the bed nucleus of the stria terminalis (BNST)), and two brainstem stress centers such as the centrally projecting Edinger-Westphal nucleus (cpEW) and dorsal raphe nucleus (DR) show age dependency in their c-Fos response. The somatosensory barrel cortex area (S1) was evaluated to test whether the age dependency is specific for stress-centers. Our results indicate that the stress-induced rise in blood CORT titer was lower in young age reflecting relatively low HPA activity. All 12 stress-related brain areas showed c-Fos response that peaked at 2 months of age. The magnitude of c-Fos immunoreactivity correlated negatively with age in seven regions (MeA, CeA, ovBNST, dlBNST, dmBNST, fuBNST and pPVN). Unexpectedly, the CeA, ovBNST and cpEW showed a considerable basal c-Fos expression in 1-month-old rats which decreased with age. The S1 showed a U-shaped age-related dynamics in contrast to the decline observed in stress centers. We conclude that the age- and brain area dependent dynamics in stress-induced neuronal activity pattern may contribute to the age dependance of the stress reactivity. Further studies are in progress to determine the neurochemical identity of neurons showing age-dependent basal and/or stress-induced c-Fos expression.

P.3.c.003 - Nigella sativa extract improved sensorimotor gating deficit on acute ketamine model of schizophrenia in rats

A traditional Japanese (Kampo) medicine, yokukansan, has long been used to treat neurosis, insomnia, and night crying and irritability in children. Recently, this medicine has reported to improve the behavioral and psychological symptoms of dementia that often become problematic in patients with Alzheimer's disease and other forms of dementia.Several animal studies have reported that yokukansan has an anxiolytic effect. However, the underlying mechanisms are not yet understood. In the present study, we investigated the effects in rats of single and repeated administrations of yokukansan on anxiety-like behaviors, stress responses, and the brain regions involved.Yokukansan dissolved in water (100 or 300 mg/kg) was administered orally to F344/N male rats 1 h before each test or for two weeks before the tests began. Locomotor activity and anxiety-related behavior in the open-field test and the elevated plus-maze test, serum corticosterone levels, and restraint stress-induced c-Fos expression in various brain regions as a marker of neuronal activation were evaluated in both the vehicle-treated and yokukansan-treated rats.A single administration of yokukansan had no effect on locomotor activity or anxiety-like behavior; however, repeated administration decreased anxiety-like behavior in a dose-dependent manner. Neither single nor repeated administration of yokukansan had an effect on the basal or stress-induced levels of serum corticosterone. For c-Fos expression, restraint stress increased the number of c-Fos-positive cells in the subdivisions of the prefrontal cortex, amygdala, and hypothalamus. Repeated administration of yokukansan decreased the stress-induced c-Fos expression in the prelimbic cortex and the basolateral and medial amygdaloid nuclei.The present study indicates that repeated oral administration of yokukansan has an anxiolytic effect and that this effect may be associated with attenuated neuronal activity in the medial prefrontal cortex and amygdala.

Differential stress induced c-Fos expression and identification of region-specific miRNA-mRNA networks in the dorsal raphe and amygdala of high-responder/low-responder rats

Chronic stress triggers a variety of physical and mental health problems, and how individuals cope with stress influences risk for emotional disorders. To investigate molecular mechanisms underlying distinct stress coping styles, we utilized rats that were selectively-bred for differences in emotionality and stress reactivity. We show that high novelty responding (HR) rats readily bury a shock probe in the defensive burying test, a measure of proactive stress coping behavior, while low novelty responding (LR) rats exhibit enhanced immobility, a measure of reactive coping. Shock exposure in the defensive burying test elicited greater activation of HR rats’ caudal dorsal raphe serotonergic cells compared to LRs, but lead to more pronounced activation throughout LRs’ amygdala (lateral, basolateral, central, and basomedial nuclei) compared to HRs. RNA-sequencing revealed 271 mRNA transcripts and 33 microRNA species that were differentially expressed in HR/LR raphe and amygdala. We mapped potential microRNA-mRNA networks by correlating and clustering mRNA and microRNA expression and identified networks that differed in either the HR/LR dorsal raphe or amygdala. A dorsal raphe network linked three microRNAs which were down-regulated in LRs (miR-206-3p, miR-3559-5p, and miR-378a-3p) to repression of genes related to microglia and immune response (Cd74, Cyth4, Nckap1l, and Rac2), the genes themselves were up-regulated in LR dorsal raphe. In the amygdala, another network linked miR-124-5p, miR-146a-5p, miR-3068-3p, miR-380-5p, miR-539-3p, and miR-7a-1-3p with repression of chromatin remodeling-related genes (Cenpk, Cenpq, Itgb3bp, and Mis18a). Overall this work highlights potential drivers of gene-networks and downstream molecular pathways within the raphe and amygdala that contribute to individual differences in stress coping styles and stress vulnerabilities.

Adolescent caffeine consumption increases adulthood anxiety-related behavior and modifies neuroendocrine signaling

Caffeine is a commonly used psychoactive substance and consumption by children and adolescents continues to rise. Here, we examine the lasting effects of adolescent caffeine consumption on anxiety-related behaviors and several neuroendocrine measures in adulthood. Adolescent male Sprague-Dawley rats consumed caffeine (0.3g/L) for 28 consecutive days from postnatal day 28 (P28) to P55. Age-matched control rats consumed water. Behavioral testing for anxiety-related behavior began in adulthood (P62) 7 days after removal of caffeine. Adolescent caffeine consumption enhanced anxiety-related behavior in an open field, social interaction test, and elevated plus maze. Similar caffeine consumption in adult rats did not alter anxiety-related behavior after caffeine removal. Characterization of neuroendocrine measures was next assessed to determine whether the changes in anxiety were associated with modifications in the HPA axis. Blood plasma levels of corticosterone (CORT) were assessed throughout the caffeine consumption procedure in adolescent rats. Adolescent caffeine consumption elevated plasma CORT 24h after initiation of caffeine consumption that normalized over the course of the 28-day consumption procedure. CORT levels were also elevated 24h after caffeine removal and remained elevated for 7 days. Despite elevated basal CORT in adult rats that consumed caffeine during adolescence, the adrenocorticotropic hormone (ACTH) and CORT response to placement on an elevated pedestal (a mild stressor) was significantly blunted. Lastly, we assessed changes in basal and stress-induced c-fos and corticotropin-releasing factor (Crf) mRNA expression in brain tissue collected at 7 days withdrawal from adolescent caffeine. Adolescent caffeine consumption increased basal c-fos mRNA in the paraventricular nucleus of the hypothalamus. Adolescent caffeine consumption had no other effects on the basal or stress-induced c-fos mRNA changes. Caffeine consumption during adolescence increased basal Crf mRNA in the central nucleus of the amygdala, but no additional effects of stress or caffeine consumption were observed in other brain regions. Together these findings suggest that adolescent caffeine consumption may increase vulnerability to psychiatric disorders including anxiety-related disorders, and this vulnerability may result from dysregulation of the neuroendocrine stress response system.

Ameliorating effects of gypenosides on chronic stress-induced anxiety disorders in mice

BackgroundEthanol extract from Gynostemma pentaphyllum (GP) shows anti-stress and anxiolytic functions in mice, and also protects dopamine neurons in 6-hydroxydopamine-lesioned rat model of Parkinson’s disease. In addition, gypenosides (the gypenoside-enriched components of GP, GPS) have a protective effect on 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced mouse model of Parkinson’s disease. In this study, the ameliorating effects of GPS on chronic stress-induced anxiety disorders in mice were investigated.MethodsMice were orally treated with GPS (100 and 200 mg/kg) once a day for 10 days, followed by exposure to electric footshock (EF) stress (0.6 mA, 1 s every 5 s, 3 min). After the final administration of either GPS, water extract of GP (GP-WX) or ethanol extract of GP (GP-EX, positive control), the behavioral tests such as elevated plus-maze, marble burying and locomotor activity tests, and the biochemical parameters including dopamine, serotonin and corticosterone levels, and c-Fos expression were examined.ResultsTreatment with GPS (100 and 200 mg/kg) increased the number of open arm entries and the time spent on open arms in elevated plus-maze which were reduced by chronic EF stress. GPS (100 and 200 mg/kg) reduced the number of marbles buried which increased by chronic EF stress. In these states, the brain levels of dopamine and serotonin decreased by chronic EF stress and they were recovered by GPS. The serum levels of corticosterone increased by chronic EF stress were also reduced by GPS (100 and 200 mg/kg). Finally, chronic EF stress-induced c-Fos expression was markedly reduced by GPS (100 and 200 mg/kg) in the brain. GPS (100 and 200 mg/kg) also showed an equivalent efficacy on anxiolytic functions, as compared with GP-EX (50 mg/kg). However, GP-WX (50 mg/kg) showed a less effect on anxiety disorders than GP-EX (50 mg/kg) and GPS (100 and 200 mg/kg).ConclusionThese results suggest that GPS (100 and 200 mg/kg) has anxiolytic effects on chronic EF stress-induced anxiety disorders by modulating dopamine and serotonin neuronal activities, c-Fos expression and corticosterone levels. GPS may serve as a phytonutrient in chronic stress-induced anxiety disorders.

Presentation of noise during acute restraint stress attenuates expression of immediate early genes and arginine vasopressin in the hypothalamic paraventricular nucleus but not corticosterone secretion in rats

The present study investigated the effect of acoustic stimulation on the activation of the hypothalamic-pituitary-adrenal (HPA) axis in rats submitted to acute restraint stress, through semi-quantitative histochemical analysis of expression of immediate early gene products (c-Fos, JunB and phosphorylated c-Jun) and arginine vasopressin (AVP) hnRNA in the paraventricular nucleus (PVN). Simultaneous presentation of white or pink noise with restraint resulted in a significant attenuation of stress-induced c-Fos and JunB expression in the dorsal body of dorsal medial parvicellular subdivision (mpdd) of the PVN, as compared with restraint without noise. However, this presentation did not change phosphorylation of c-Jun and the plasma corticosterone level. Moreover, white noise presentation during restraint led to a reduction in the number of c-Fos- or JunB-expressing corticotropin-releasing hormone (CRH) neurons and the number of neurons expressing AVP hnRNA in the mpdd. Dual-histochemical labeling revealed co-expression of c-Fos and JunB, as well as JunB and AVP hnRNA in mpdd neurons. These data suggest that acoustic stimuli have an attenuation effect on the restraint-induced activation of neuroendocrine CRH neurons, resulting in the reduction in AVP production as an adaptation of HPA axis to repeated stress.

Effects of gypenosides on chronic stress-induced anxiety in mice

The objective of present study was to investigate the effects of gypenosides (GPS), the gypenosides-enriched components of Gynostemma pentaphyllum, on chronic stress-induced anxiety in mice. Animals were treated with GPS (50, 100 and 200 mg/kg, oral) for 10 days before the exposure to chronic electric footshock (EF) stress (intensity 2 mA, interval 10 s and duration 3 min). The chronic stressed-group showed anxiety disorders in elevated plus maze compared with control group. However, the treatment of GPS ameliorated the anxiety disorders induced by chronic EF stress. The spontaneous locomotor activities were decreased by ca. 18.1% (P < 0.05) by chronic EF stress compared with control group (n = 10). The spontaneous locomotor activities decreased were increased by ca. 9.4%, 15.3% (P < 0.05) and 15.9% (P < 0.05) by treatment with GPS (50, 100 and 200 mg/kg). Furthermore, the levels of dopamine and serotonin in the brain were decreased by ca. 26.6% (P < 0.05) and 25.1% (P < 0.05) after being exposed to chronic EF stress compared with control group (n = 10). The levels of dopamine and serotonin were inceased by ca. 11.4%, 18.0% (P < 0.05), 16.6% (P < 0.05) and 9.1%, 17.6% (P < 0.05), 15.7% (P < 0.05), respectively, by GPS (50, 100 and 200 mg/kg) compared with chronic EF stressed-group (n = 10). Finally, chronic EF stress-induced c-Fos expression in the brain was markedly increase by ca. 188% (P < 0.01) compared with control group. However, the number of c-Fos-immunoreactive cells were significantly decreased by ca. 19.4% (P < 0.05), 30.2% (P < 0.01) and 24.3% (P < 0.05) by treatment with GPS (50, 100 and 200 mg/kg), compared with chronic EF stressed-group. In addition, the anxiolytic functions of GPS were compared with GP-EX, ethanol extracts from Gynostemma pentaphyllum. These results suggest that GPS have anxiolytic effects on chronic EF stress-induced anxiety disorders and GPS may serve as an herbal drug in chronic stress-induced anxiety disorders.

Elevated corticosterone in the dorsal hindbrain increases plasma norepinephrine and neuropeptide Y, and recruits a vasopressin response to stress.

Repeated stress and chronically elevated glucocorticoids cause exaggerated cardiovascular responses to novel stress, elevations in baseline blood pressure, and increased risk for cardiovascular disease. We hypothesized that elevated corticosterone (Cort) within the dorsal hindbrain (DHB) would: 1) enhance arterial pressure and neuroendocrine responses to novel and repeated restraint stress, 2) increase c-Fos expression in regions of the brain involved in sympathetic stimulation during stress, and 3) recruit a vasopressin-mediated blood pressure response to acute stress. Small pellets made of 10% Cort were implanted on the surface of the DHB in male Sprague-Dawley rats. Blood pressure was measured by radiotelemetry. Cort concentration was increased in the DHB in Cort-treated compared with Sham-treated rats (60 ± 15 vs. 14 ± 2 ng Cort/g of tissue, P < 0.05). DHB Cort significantly increased the integrated arterial pressure response to 60 min of restraint stress on days 6, 13, and 14 following pellet implantation (e.g., 731 ± 170 vs. 1,204 ± 68 mmHg/60 min in Sham- vs. Cort-treated rats, day 6, P < 0.05). Cort also increased baseline blood pressure by day 15 (99 ± 2 vs. 108 ± 3 mmHg for Sham- vs. Cort-treated rats, P < 0.05) and elevated baseline plasma norepinephrine and neuropeptide Y concentrations. Cort significantly enhanced stress-induced c-Fos expression in vasopressin-expressing neurons in the paraventricular nucleus of the hypothalamus, and blockade of peripheral vasopressin V1 receptors attenuated the effect of DHB Cort to enhance the blood pressure response to restraint. These data indicate that glucocorticoids act within the DHB to produce some of the adverse cardiovascular consequences of chronic stress, in part, by a peripheral vasopressin-dependent mechanism.

Increased depression-like behaviors with dysfunctions in the stress axis and the reward center by free access to highly palatable food

This study was conducted to examine the behavioral consequences of unlimited consumption of highly palatable food (HPF) and investigate its underlying neural mechanisms. Male Sprague–Dawley rats had free access to chocolate cookie rich in fat (HPF) in addition to ad libitum chow and the control group received chow only. Rats were subjected to behavioral tests during the 2nd week of food condition; i.e. ambulatory activity test on the 8th, elevated plus maze test (EPM) on the 10th and forced swim test (FST) on the 14th day of food condition. After 8days of food condition, another group of rats were placed in a restraint box and tail bloods were collected at 0, 20, 60, and 120 time points during 2h of restraint period, used for the plasma corticosterone assay. At the end of restraint session, rats were sacrificed and the tissue sections of the nucleus accumbens (NAc) were processed for c-Fos immunohistochemistry. Ambulatory activities and the scores of EPM were not significantly affected by unlimited cookie consumption. However, immobility duration during FST was increased, and swim decreased, in the rats received free cookie access compared with control rats. Stress-induced corticosterone increase was exaggerated in cookie-fed rats, while the stress-induced c-Fos expression in the NAc was blunted, compared to control rats. Results suggest that free access to HPF may lead to the development of depression-like behaviors in rats, likely in relation with dysfunctions in the hypothalamic–pituitary–adrenal axis and the reward center.

Brain region-specific reduction in c-Fos expression associated with an anxiolytic effect of yokukansan in rats

Ethnopharmacological relevanceA traditional Japanese (Kampo) medicine, yokukansan, has long been used to treat neurosis, insomnia, and night crying and irritability in children. Recently, this medicine has reported to improve the behavioral and psychological symptoms of dementia that often become problematic in patients with Alzheimer's disease and other forms of dementia. Aim of the studySeveral animal studies have reported that yokukansan has an anxiolytic effect. However, the underlying mechanisms are not yet understood. In the present study, we investigated the effects in rats of single and repeated administrations of yokukansan on anxiety-like behaviors, stress responses, and the brain regions involved. Materials and methodsYokukansan dissolved in water (100 or 300mg/kg) was administered orally to F344/N male rats 1h before each test or for two weeks before the tests began. Locomotor activity and anxiety-related behavior in the open-field test and the elevated plus-maze test, serum corticosterone levels, and restraint stress-induced c-Fos expression in various brain regions as a marker of neuronal activation were evaluated in both the vehicle-treated and yokukansan-treated rats. ResultsA single administration of yokukansan had no effect on locomotor activity or anxiety-like behavior; however, repeated administration decreased anxiety-like behavior in a dose-dependent manner. Neither single nor repeated administration of yokukansan had an effect on the basal or stress-induced levels of serum corticosterone. For c-Fos expression, restraint stress increased the number of c-Fos-positive cells in the subdivisions of the prefrontal cortex, amygdala, and hypothalamus. Repeated administration of yokukansan decreased the stress-induced c-Fos expression in the prelimbic cortex and the basolateral and medial amygdaloid nuclei. ConclusionsThe present study indicates that repeated oral administration of yokukansan has an anxiolytic effect and that this effect may be associated with attenuated neuronal activity in the medial prefrontal cortex and amygdala.

Environmental Enrichment and Gut Inflammation Modify Stress-Induced c-Fos Expression in the Mouse Corticolimbic System

Environmental enrichment (EE) has a beneficial effect on rodent behaviour, neuronal plasticity and brain function. Although it may also improve stress coping, it is not known whether EE influences the brain response to an external (psychological) stressor such as water avoidance stress (WAS) or an internal (systemic) stressor such as gastrointestinal inflammation. This study hence explored whether EE modifies WAS-induced activation of the mouse corticolimbic system and whether this stress response is altered by gastritis or colitis. Male C67BL/6N mice were housed under standard or enriched environment for 9 weeks, after which they were subjected to a 1-week treatment with oral iodoacetamide to induce gastritis or oral dextran sulfate sodium to induce colitis. Following exposure to WAS the expression of c-Fos, a marker of neuronal activation, was measured by immunocytochemistry. EE aggravated experimentally induced colitis, but not gastritis, as shown by an increase in the disease activity score and the colonic myeloperoxidase content. In the brain, EE enhanced the WAS-induced activation of the dentate gyrus and unmasked an inhibitory effect of gastritis and colitis on WAS-evoked c-Fos expression within this part of the hippocampus. Conversely, EE inhibited the WAS-evoked activation of the central amygdala and prevented the inhibitory effect of gastritis and colitis on WAS-evoked c-Fos expression in this region. EE, in addition, blunted the WAS-induced activation of the infralimbic cortex and attenuated the inhibitory effect of gastritis and colitis on WAS-evoked c-Fos expression in this area. These data reveal that EE has a region-specific effect on stress-induced c-Fos expression in the corticolimbic system, which is likely to improve stress resilience. The response of the prefrontal cortex – amygdala – hippocampus circuitry to psychological stress is also modified by the systemic stress of gut inflammation, and this interaction between external and internal stressors is modulated by the housing environment.

Environmental enrichment and visceral inflammation regulate stress-induced c-Fos and NPY expression within the dentate gyrus

Background The dentate gyrus, a part of the hippocampal formation, is an important brain region regulating the central response to stress. Given that stress resilience depends on genetics and adaptive processes within the brain, the question arises as to whether the acute stress response is modified by chronic environmental or pathological conditions. Therefore we investigated, on the one hand, whether environmental enrichment (EE), a housing condition suggested to promote stress resilience, alters the acute stress response. On the other hand, we assessed whether visceral inflammation, known to be exacerbated by stress, also has an impact on the central stress response.

Nicotine modulates multiple regions in the limbic stress network regulating activation of hypophysiotrophic neurons in hypothalamic paraventricular nucleus

Nicotine intake affects CNS responses to stressors. We reported that nicotine self-administration (SA) augmented the hypothalamo-pituitary-adrenal (HPA) stress response, in part because of the altered neurotransmission and neuropeptide expression within hypothalamic paraventricular nucleus (PVN). Limbic-PVN interactions involving medial prefrontal cortex, amygdala, and bed nucleus of the stria terminalis (BST) greatly impact the HPA stress response. Therefore, we investigated the effects of nicotine SA on stress-induced neuronal activation in limbic-PVN network, using c-Fos protein immunohistochemistry and retrograde tracing. Nicotine decreased stress-induced c-Fos in prelimbic cortex (PrL), anteroventral BST (avBST), and peri-PVN, but increased c-Fos induction in medial amygdala (MeA), locus coeruleus, and PVN. Fluoro-gold (FG) was injected into avBST or PVN, as GABAergic neurons in avBST projecting to PVN corticotrophin-releasing factor neurons relay information from both PrL glutamatergic and MeA GABAergic neurons. The stress-induced c-Fos expression in retrograde-labeled FG+ neurons was decreased in PrL by nicotine, but increased in MeA, and also reduced in avBST. Therefore, within limbic-PVN network, nicotine SA exerts selective regional effects on neuronal activation by stress. These findings expand the mechanistic framework by demonstrating altered limbic-BST-PVN interactions underlying the disinhibition of PVN corticotrophin-releasing factor neurons, an essential component of the amplified HPA response to stress by nicotine.

The effect of auditory stimulation on restraint stress-induced c-Fos expression in the paraventricular nucleus of rats

The conditioned place preference (CPP) paradigm is widely used when examining the reinforcing effects of drugs. Some previous studies have shown that an acute stressor, such as restraint could also induce CPP. Although the modulating effects of stress hormones on various forms of learning are well known, the finding that a stressor has a potentially direct role in the reinforcement mechanism is novel. This study focused on the function of stress hormones in restraint-induced CPP in Wistar rats administered agonist or antagonist of 2 critical stress hormones prior to conditioning. Results showed that peripheral applications of corticosterone (CORT, 1, 3, 5, and 10 mg/kg, subcutaneously) failed to induce CPP. Furthermore, a glucocorticoid (GC) antagonist (mifepristone, 10, 40, or 100 mg/kg, sc) failed to block the restraint-induced CPP. Intracerebroventricular injection of a selective corticotropin-releasing factor receptor 1 (CRFR1) antagonist antalarmin (1 μg/5 μl), on the contrary, completely blocked the restraint-induced CPP. We concluded that CRFR1 plays an essential role in the neural mechanism of restraint-induced CPP. Negative feedback of CORT from peripheral sources may not be involved in this phenomenon.